Spataro A C, Morgan H R, Bosmann H B
J Cell Sci. 1976 Jul;21(2):407-13. doi: 10.1242/jcs.21.2.407.
The proteolytic activities of normal, Schmidt-Ruppin Rous sarcoma virus (SR-RSV) transformed, and infected (RAV) chick embryo fibroblasts (CEF) have been measured by a highly sensitive technique using 3H-acetylated haemoglobin as a substrate. When all 3 types of CEF cells were maintained in serumless media, no differences were detected in the amount of pH 3-4 protease activity released into the media over a 24-h period, and only negligible amounts of pH 7-6 proteolytic activity were found. When normal, transformed, and infected cells were maintained in serumless media and later incubated with 3H-acetylate haemoglobin, a significant proteolysis of the haemoglobin, a 6-fold increase compared to the normal CEF cells, was associated only with plates containing SR-RSV-CEF cells. A fluorescent assay for peptides confirmed that SR-RSV-CEF cells have increased cell-associated proteolytic activity. The net surface charge of the transformed CEF cells was unchanged by maintenance in serumless media but the net surface negativity of the normal and RAV-CEF cells was significantly increased by incubation in media minus serum for 24 h. This suggests that normal CEF cells, maintained in media plus serum, have a substance masking their surface charge which is absent from the surface of transformed cells, possibly because of proteolytic degradation.
采用以³H - 乙酰化血红蛋白为底物的高灵敏度技术，测定了正常、施密特 - 鲁平劳氏肉瘤病毒（SR - RSV）转化以及感染劳氏禽肉瘤病毒（RAV）的鸡胚成纤维细胞（CEF）的蛋白水解活性。当所有这3种类型的CEF细胞在无血清培养基中培养时，在24小时内释放到培养基中的pH 3 - 4蛋白酶活性量未检测到差异，且仅发现极少量的pH 7 - 6蛋白水解活性。当正常、转化和感染的细胞在无血清培养基中培养，随后与³H - 乙酰化血红蛋白一起孵育时，血红蛋白的显著蛋白水解（与正常CEF细胞相比增加了6倍）仅与含有SR - RSV - CEF细胞的平板相关。肽的荧光测定证实，SR - RSV - CEF细胞具有增加的细胞相关蛋白水解活性。转化的CEF细胞的净表面电荷在无血清培养基中培养时未改变，但正常和RAV - CEF细胞的净表面负性在无血清培养基中孵育24小时后显著增加。这表明，在含血清培养基中培养的正常CEF细胞有一种掩盖其表面电荷的物质，而转化细胞表面不存在这种物质，这可能是由于蛋白水解降解所致。
